and B may fade or disappear within a few minutes, record reaction as soon as color appears. If no color develops, test for presence of nitrate by adding small amount of zinc dust. If color develops, nitrate has not been reduced.

Nitrate reduction test for enteropathogenic E. coli. To 3 ml of 18- 24 h culture in indole-nitrite medium, add 2 drops each of reagents A and B.Red-violet color indicates that nitrate has been reduced to nitrite. Check negative tests by adding small amount of zinc dust; if red-violet color does not appear, nitrate has been reduced.

D. Alternative test reagents. 5-Amino-2-naphthylene sulfonic acid (Cleve's acid) and N,N-dimethyl-1-naphthylamine have been recommended as substitutes for preparation of reagent B. Absolute ethanol may be substituted for acetic acid in reagent C. However, comparative evaluations should be conducted before substitution of these alternative reagents.

This is the preferred reagent. Use freshly prepared. However, reagent can be used up to 7 days if stored in a dark glass bottle under refrigeration.Apply freshly prepared solution directly to young culture (24 h) on either agar plate or slant. Oxidase-positive colonies develop a pink color and progressively turn dark purple. If cultures are to be preserved, complete the transfer from plates to which